Abstract

The Ccr4‐Not complex is a multisubunit complex present in all eukaryotes that contributes to regulate gene expression at all steps, from production of messenger RNAs (mRNAs) in the nucleus to their degradation in the cytoplasm. In the nucleus it influences the post‐translational modifications of the chromatin template that has to be remodeled for transcription, it is present at sites of transcription and associates with transcription factors as well as with the elongating polymerase, it interacts with the factors that prepare the new transcript for export to the cytoplasm and finally is important for nuclear quality control and influences mRNA export. In the cytoplasm it is present in polysomes where mRNAs are translated and in RNA granules where mRNAs will be redirected upon inhibition of translation. It influences mRNA translatability, and is needed during translation, on one hand for co‐translational protein interactions and on the other hand to preserve translation that stalls. It is one of the relevant players during co‐translational quality control. It also interacts with factors that will repress translation or induce mRNA decapping when recruited to the translating template. Finally, Ccr4‐Not carries deadenylating enzymes and is a key player in mRNA decay, generic mRNA decay that follows normal translation termination, co‐translational mRNA decay of transcripts on which the ribosomes stall durably or which carry a non‐sense mutation and finally mRNA decay that is induced by external signaling for a change in genetic programming. Ccr4‐Not is a master regulator of eukaryotic gene expression. WIREs RNA 2016, 7:438–454. doi: 10.1002/wrna.1332 This article is categorized under: Translation > Translation Regulation RNA Turnover and Surveillance > Turnover/Surveillance Mechanisms RNA Turnover and Surveillance > Regulation of RNA Stability

Images

Linear and cartoon representation of the Ccr4‐Not complex subunits. (a) The Not1 core subunit is depicted with the domains characterized as docking sites for other core subunits. The names chosen for the core subunits are those described in Table . DEDD and EEP refer to the signature domains of the catalytic deadenylase subunits and RING to the signature domain of the E3 ligase subunit. NOT box is a domain of homology shared by several Not proteins and RRM is a putative RNA recognition motif in Not4. Adapted from Ref to include the docking of Not4 defined in Ref . The C‐terminal domain of Not4 binds the side surface of the first HEAT‐repeat unit of the Not1 C‐terminal domain, while Not2 together with the C‐terminal domain of Not5 bind the top and the bottom surfaces. Hence the interactions occur at largely separate surfaces of the Not1 C‐terminal domain. (b) Cartoon representation of the L‐shape of the Ccr4‐Not complex defined by electron microscopy with the expected position of the core subunits on the Not1 scaffold. Where Not3 and Caf130 dock onto Not1 is unknown, so we placed Not3 in contact with Not2 and Not5 because of common mutant phenotypes and the yeast specific Caf130 at the N‐terminus of Not1 where Not10 and Not11 bind in flies.

Ccr4‐Not connects the nuclear and cytoplasmic phases of gene expression. Adapted from Ref . Ccr4‐Not is present at sites of transcription by interaction with transcription factors (TFs) and with the elongating RNAPII via Rpb4. It might associate with newly produced mRNAs and be exported to the nucleus with the mRNPs. In the cytoplasm it will contribute to regulate translation and bring about mRNA degradation. But it will also promote assembly of new RNAPII that can re‐enter the nucleus to start new rounds of transcription.

The Ccr4‐Not is the major eukaryotic deadenylase. (a) Ccr4‐Not can be recruited to mRNAs via an interaction between Caf1 and Tob1 itself binding to PABP. This can cause a generalized deadenylation of mRNAs upon Tob1 expression. (b) During NMD in verterbrates, phosphorylated Upf1 can recruit the Smg5, 6, and 7 proteins. Smg6 is an endonuclease that acts redundantly to the Ccr4‐Not complex recruited by the Smg5/7 heterodimer, via the interaction of Smg7 and Pop2, to bring about mRNA decay. (c) RBPs can tether the Ccr4‐Not complex to target mRNAs and lead to deadenylation probably after normal translation termination. (d) The micro‐RNA machinery can tether Ccr4‐Not and bring about mRNA deadenylation after normal translation termination.

The Ccr4‐Not complex plays many roles during the translation process. (a) Not5 is needed for the presence of the R2TP co‐chaperone at polysomes producing Rpb1 to form a soluble assembly‐competent Rpb1 including Hsp90 and the Rpb4, 5, 6, 7, and 9 subunits, that will form RNAPII. (b) Not4 is needed probably co‐translationally for the interaction of the Ecm29 chaperone with regulatory particle (RP) proteasome subunits and proteasome integrity. CP: core particle. (c) Not4 is a relevant player during co‐translational quality control. It has reported positive and negative effects on translation initiation, the ability to ubiquitinate stalled peptide or finally to initiate mRNA decay by Ccr4‐Not dependent deadenylation. (d) Upon recruitment by an RNA binding protein (RBP) Ccr4‐Not mediates translational repression in a deadenylation‐independent manner via the recruitment of the 4E‐T protein. 4E‐T interacts with eIF4E and decapping components associating with the 3′ and 5′ ends of mRNAs such as the LSM, PAT1, and DDX6 proteins. DDX6 can also directly interact with Not1. (e) Cartoon with a model for both positive and negative effects of Not4 on translation.

Cartoon depicting the different interactions between Ccr4‐Not and components contributing to mRNA export and nuclear quality control. Ccr4‐Not interacts with Hpr1 and Mft1 or the THO/TREX complex that includes Sub2, with the nuclear poly(A) binding protein Nab2, with Mlp1, and is necessary for the interaction of the Rrp6 exonuclease with its co‐factor the Mtr4 RNA helicase. Some other components that participate in export such as Mex67, Mtr2, Yra1, and Mlp1 are included. NPC: nuclear pore complex.

Representation of the roles of Ccr4‐Not in transcription elongation. Adapted from Refs . On the right is depicted RNAPII that encounters a block in transcription during the productive elongation phase and that backtracks. On the upper left panel is depicted reiterative transcript binding and release by Ccr4–Not that causes realignment of the 3′ end of the RNA into the active site and promotes the resumption of elongation by RNAPII. On the lower left panel is depicted that Ccr4‐Not increases the recruitment of TFIIS to backtracked RNAPII complexes. This enhances the cleavage of the displaced transcript in backtracked RNAPII by TFIIS, allowing elongation to resume.